title: DASE11-2
description: Protocols: For RNA sequencing, we pooled the basal-most 2 cm sections of 25 silk strands from the mid-base region of four individual ears to obtain four biological replicates. Silk bases were collected 3-, 7-, 11- and 15 days after silk emergence (DASE). greenhouse Total RNA from silk material was extracted and purified with Spectrum™ Plant Total RNA Kit (Sigma-Aldrich, Belgium). RNA concentration and purity were quantified by Nanodrop spectrophotometry (Nanodrop Technologies, Wilmington, DE, USA), and RNA integrity (RIN) was confirmed by an Agilent 2100 Bioanalyzer (Agilent Technologies, Palo Alto, CA, USA).. Library preparation was done using the TruSeq RNA Sample Preparation Kit v2 (Illumina). In brief, poly(A)-containing mRNA molecules were reverse transcribed, double-stranded cDNA was generated, and adapters were ligated. After quality control using the 2100 Bioanalyzer, clusters were generated through amplification using the TruSeq PE Cluster Kit v3-cBot-HS kit (Illumina) followed by sequencing on Illumina NextSeq 500 with the TruSeq SBS Kit v3-HS (Illumina).